The present invention relates generally to a process for crystallizing egg white lysozyme, and more particularly to a process for crystallizing egg white lysozyme by which crystallized egg white lysozyme can be collected more rapidly than heretofore.
Heretofore, the so-called "crude lysozyme" extracted from egg white by adsorption or salting out has been purified ordinarily by adjusting the pH of an aqueous sodium chloride solution containing the crude lysozyme dissolved therein to a value of the order of 9.5 so that crystalline lysozyme will be formed by isoelectric precipitation. Ordinarily, in this process, the aqueous sodium chloride solution is cooled to a temperature of the order of 0.degree. C. to 5.degree. C. and then a small quantity of a seed crystal is added, or a small quantity of a seed crystal is added and immediately thereafter the solution is cooled to a temperature of the same order thereby to facilitate the formation of crystalline lysozyme.
However, when 1 liter, for example, of an aqueous sodium chloride solution opacified or clouded with crystallized lysozyme obtained after about 10 hours of crystallization in accordance with the process described above is filtered through Toyo Filter Paper No. 2 having a filter area of 300 cm.sup.2 under a gauge pressure of 1.5 kg/cm.sup.2 to collect the lysozyme crystals, more than 20 minutes are required for the filtration presumably because the filter paper clogs. Thus, this crystallization process is accompanied by the problem of an excessively long period of time required for the subsequent filtration operation.
We have continued extensive research in order not only to solve this problem encountered in the purification of crude lysozyme mentioned above but also to overcome a similar problem encountered in the crystallization of lysozyme by a salt concentration adjustment from a common salt solution containing lysozyme dissolved therein, such as an eluate obtained by eluting lysozyme adsorbed on any type of resin in accordance with the conventional adsorption method. That is, an object of our study was to provide a novel process for crystallizing lysozyme by which crystallized lysozyme can be collected more rapidly or, more particularly, by which filtration can be simplified.
As a result, it has been found that, by adding a seed crystal to a salt solution containing lysozyme, maintaining the solution at about 10.degree. C. to 28.degree. C. for a certain period of time, and then cooling the resulting solution, the subsequent filtration operation can be completed within an extremely short period of time, that is, within only a few minutes.